Apparatus for testing, assessment, and maintenance of harvested hearts for transplanting

ABSTRACT

An apparatus, a system, and methods for receiving, perfusing and maintaining and assessing excised donor heart physiological functionality. The system generally comprises an apparatus for receiving and holding an excised heart that is interconnected with: (i) a perfusate-processing system, (ii) a bi-directional perfusate pumping system, (iii) flow sensors for monitoring the flow of perfusate to and from an installed heart&#39;s aorta, pulmonary artery, pulmonary vein, and vena cava, (iv) an ECG apparatus interconnectable with the installed heart, and (v) probes interconnecting the installed heart with instruments for monitoring the heart&#39;s physiogical functionality using load independent indices and load dependent indices.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.14/372,909, filed on Jul. 17, 2014, entitled “APPARATUS FOR TESTING,ASSESSMENT, AND MAINTENANCE OF HARVESTED HEARTS FOR TRANSPLANTING”,which is a national filing of PCT International Application No.PCT/CA2013/000031, filed Jan. 17, 2013, entitled “APPARATUS FOR TESTING,ASSESSMENT, AND MAINTENANCE OF HARVESTED HEARTS FOR TRANSPLANTING”,which claims benefit of, and priority from, U.S. Provisional PatentApplication No. 61/587,452, filed Jan. 17, 2012, the entire contents ofeach of which are incorporated herein by reference.

FIELD OF THE INVENTION

The present invention pertains to apparatus, systems, and methods for exvivo perfusion and maintenance of harvested donor hearts, and moreparticularly, to pre-transplant assessment of harvested donor hearts fortheir suitability for transplantation.

BACKGROUND OF THE INVENTION

Heart failure affects 10% of North Americans and is the leading hospitaldischarge diagnosis. The diagnosis of heart failure is accompanied by asurvival outlook that is comparable to a major cancer. There are limitedrehabilitation options available to patients who are suffering withheart failure, and few strategies actually re-power the heart. Cardiactransplantation remains the gold-standard therapeutic intervention forpatients with end-stage heart failure, with an increasing number ofindividuals being added to the transplant wait list every year. However,wider application of this life-preserving intervention is limited by theavailability of donors. Data from the International Society of Heart andLung Transplantation Registry shows that cardiac transplantation is inprogressive decline in suitable donors (2007, Overall Heart and AdultHeart Transplantation Statistics). Two hundred and fifty eight Canadianshave died during the last decade (2000-2010; Heart and Stroke Foundationof Canada) while waiting for heart transplantation. Similarly, in theUnited States, 304 patients died in 2010 alone while waiting for hearttransplantation (Organ Procurement and Transplantation Network, US Dept.of Health & Human Services). This phenomenon is primarily due to ashortage of suitable organ donors, and is being experienced across theglobe.

Time is of the essence for removal of a heart from a donor and itssuccessful transplantation into a recipient. The following principlesgenerally apply for optimal donor heart preservation for the period oftime between removal from the donor and transplantation: (i)minimization of cell swelling and edema, (ii) prevention ofintracellular acidosis, (iii) prevention of injury caused by oxygen freeradicals, and (iv) provision of substrate for regeneration ofhigh-energy phosphate compounds and ATP during reperfusion. The two mainsources of donor hearts for transplantation are breathing patients whohave suffered irreversible loss of brain function as a result of blunthead trauma or intracerebral hemorrhage and are classified as“brainstem-dead” donors, and patients who have suffered circulatorydeath and are referred to as “non-heart-beating” donors.

Brainstem-dead organ donors can be maintained under artificialrespiration for extended periods of time to provide relative hemodynamicstability up throughout their bodies until the point of organ retrieval.Therefore, cardiac perfusion is uncompromised and organ functionality istheoretically maintained. However, brainstem death itself can profoundlyaffect cardiac function. The humoral response to brainstem death ischaracterized by a marked rise in circulating catecholaminesPhysiological responses to this “catecholamine storm” includevasoconstriction, hypertension and tachycardia, all of which increasemyocardial oxygen demand. In the coronary circulation significantincreased levels of catecholamine circulating throughout the vascularsystem induce vasoconstriction which in turn, compromises myocardialoxygen supply and can lead to subendocardial ischemia. This imbalancebetween myocardial oxygen supply and demand is one factor implicated inthe impairment of cardiac function observed following brainstem death(Halejcio-Delophont et al., 1998, Increase in myocardial interstitialadenosine and net lactate production in brain-dead pigs; an in vivomicrodialysis study. Transplantation 66(10):1278-1284;Halejcio-Delophont et al, 1998, Consequences of brain death on coronaryblood flow and myocardial metabolism. Transplant Proc. 30(6):2840-2841).Structural myocardial damage occurring after brainstem death ischaracterized by myocytolysis, contraction band necrosis,sub-endocardial hemorrhage, edema and interstitial mononuclear cellinfiltration (Baroldi et al., 1997, Type and extent of myocardial injuryrelated to brain damage and its significance in heart transplantation: amorphometric study. J. Heart Lung Transplant 16(10):994-1000). In spiteof no direct cardiac insult, brainstem-dead donors often exhibit reducedcardiac function and the current views are that only 40% of hearts canbe recovered from this donor population for transplantation.

Well-defined criteria have been developed for harvesting organs fortransplantation from non-heart-beating donors (Kootstra et al., 1995,Categories of non-heart-beating donors. Transplant Proc.27(5):2893-2894; Bos, 2005, Ethical and legal issues innon-heart-beating organ donation. Transplantation, 2005. 79(9): p.1143-1147). Non-heart-beating donors have minimal brain function but donot meet the criteria for brainstem death and therefore, cannot belegally declared brainstem dead. When it is clear that there is no hopefor meaningful recovery of the patient, the physicians and family mustbe in agreement to withdraw supportive measures. Up to this point incare, non-heart-beating patients are often supported with mechanicalventilation as well as intravenous inotropic or vasopressor medication.However, only those with single system organ failure (neurologic system)can be considered for organ donation. Withdrawal of life support, mostcommonly the cessation of mechanical ventilation, is followed by anoxiccardiac arrest after which, the patient must remain asystolic for fiveminutes before organ procurement is allowed. Consequently,non-heart-beating donors are necessarily exposed to variable periods ofwarm ischemia after cardiac arrest which may result in various degreesof organ damage. However, provided that the time duration of warmischemia is not excessive, many types organs, i.e., kidneys, livers, andlungs, harvested from non-heart-beating donors are able to recoverfunction after transplantation with success rates that approximate thosefor transplanted organs from brainstem-dead beating donors.

Numerous perfusion apparatus, systems and methods have been developedfor ex vivo maintenance and transportation of harvested organs. Mostemploy hypothermic conditions to reduce organ metabolism, lower organenergy requirements, delay the depletion of high energy phosphatereserves, delay the accumulation of lactic acid, and retardmorphological and functional deteriorations associated with disruptionof oxygenated blood supply. Harvested organs are generally perfused inthese systems with preservative solutions comprising antioxidants andpyruvate under low temperatures to maintain their physiologicalfunctionality. However, it has been found that increasing amounts offree radicals and catalytic enzymes are produced during extendedmaintenance of harvested organs in pulsing pressurized hypothermicsystems. Fluctuating perfusion pressures in such systems can damage theorgans by washing off their vascular endothelial lining and traumatizethe underlying tissues. Furthermore, the harvested organs will eluteincreasing amounts of intracellular, endothelial and membraneconstituents resulting in their further physiological debilitation.

The short-comings of hypothermic apparatus, systems and methods havebeen recognized by those skilled in these arts, and alternativeapparatus, systems and methods have been developed for preservation andmaintenance of harvested organs at temperatures in the range of about25° C. to about 35° C., commonly referred to as “normothermic”temperatures. Normothermic systems typically use perfusates based on theViaspan formulation supplemented with one or more of serum albumin as asource of protein and colloid, trace elements to potentiate viabilityand cellular function, pyruvate and adenosine for oxidativephosphorylation support, transferrin as an attachment factor; insulinand sugars for metabolic support, glutathione to scavenge toxic freeradicals as well as a source of impermeant, cyclodextrin as a source ofimpermeant, scavenger, and potentiator of cell attachment and growthfactors, a high Mg⁺⁺ concentration for microvessel metabolism support,mucopolysaccharides for growth factor potentiation and hemostasis, andendothelial growth factors (Viaspan comprises potassium lactobionate,KH₂PO₄, MgSO₄, raffinose, adenosine, glutathione, allopurinol, andhydroxyethyl starch). Other normothermic perfusation solutions have beendeveloped and used (Muhlbacher et al., 1999, Preservation solutions fortransplantation. Transplant Proc. 31(5):2069-2070). While harvestedkidneys and livers can be maintained beyond twelve hours in normothermicsystems, it has become apparent that normothermic bathing, andmaintenance of harvested hearts by pulsed perfusion beyond 12 hoursresults in deterioration and irreversible debilitation of the hearts'physiological functionality. Another disadvantage of using normothermiccontinuous pulsed perfusion systems for maintenance of harvested heartsis the time required to excise the heart from a donor, mount it into thenormothermic perfusion system and then initiate and stabilize theperfusion process. After the excised heart has been stabilized, itsphysiological functionality is determined and if transplantationcriteria are met, then the excised heart is transported as quickly aspossible to a transplant facility.

SUMMARY OF THE INVENTION

The present disclosure pertains to an apparatus, a system, and methodsfor receiving, perfusion, maintenance and pre-transplant assessment ofthe physiological functionality of an excised donor heart. The systemgenerally comprises an apparatus for receiving and holding an excisedheart that is interconnected with: (i) a perfusate-processing system,(ii) a bi-directional perfusate pumping system, (iii) flow sensors formonitoring the flow of perfusate to and from an installed heart's aorta,pulmonary artery, pulmonary vein, and vena cava, (iv) an ECG apparatusinterconnectable with the installed heart, and (v) probesinterconnecting the installed heart with instruments for monitoring theheart's physiological functionality using load independent indices andload dependent indices.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention will be described in conjunction with reference tothe following drawings in which:

FIG. 1 is a schematic illustration of an exemplary testing, assessment,and maintenance apparatus for harvested donor hearts, according to oneembodiment of the present disclosure;

FIG. 2 is a perspective view showing the front of an exemplary excisedheart connected to the exemplary apparatus illustrated in FIG. 1;

FIG. 3 is a perspective view showing the back of the exemplary excisedheart connected to the exemplary apparatus;

FIG. 4 is a chart showing the effects of increasing pump pressure (RPM)on the following flow pressures of perfusate through an excised heartmounted into the exemplary apparatus: diastolic blood pressure (dbp),left artrial pressure (lap), flow rate of perfusate into the left atrium(flow), systolic blood pressure (sbp), and mean blood pressure (mbp);

FIG. 5 is a chart showing the effects of dopamine infusion concurrentincreasing pump pressure (RPM) on the following flow pressures ofperfusate through the excised heart: diastolic blood pressure (dbp),left artrial pressure (lap), flow rate of perfusate into the left atrium(flow), systolic blood pressure (sbp), and mean blood pressure (mbp);

FIG. 6 is a chart showing a family of pressure/volume loops obtainedfrom the left ventricle during the study of the effects of increasingperfusate flow pressures shown in FIG. 5;

FIG. 7 is a chart showing a family of pressure/volume loops obtainedfrom the left ventricle during the study of the effects of dopamineinfusion concurrent with increasing perfusate flow pressures shown inFIG. 5;

FIG. 8 is a chart showing a family of pressure/volume loops obtainedfrom the right ventricle during the study of the effects of increasingperfusate flow pressures shown in FIG. 5;

FIG. 9 is a chart showing a family of pressure/volume loops obtainedfrom the right ventricle during the study of the effects of dopamineinfusion concurrent with increasing perfusate flow pressures shown inFIG. 5;

FIG. 10 is a chart showing the changes in lactate concentrations in theexcised heart during the study shown in FIG. 4;

FIG. 11 is a chart showing the changes in the pH in the excised heartduring the study shown in FIG. 4; and

FIG. 12 is a schematic illustration of an exemplary testing, assessment,and maintenance apparatus for harvested donor hearts according toanother embodiment of the present disclosure.

DESCRIPTION OF THE INVENTION

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. In order that the inventionherein described may be fully understood, the following terms anddefinitions are provided herein.

The word “comprise” or variations such as “comprises” or “comprising”will be understood to imply the inclusion of a stated integer or groupsof integers but not the exclusion of any other integer or group ofintegers.

The term “about” or “approximately” means within 20%, preferably within10%, and more preferably within 5% of a given value or range.

The term “afterload” means the mean tension produced by a chamber of theheart in order to contract. It can also be considered as the ‘load’ thatthe heart must eject blood against. Afterload is therefore a consequenceof aortic large vessel compliance, wave reflection and small vesselresistance (left ventricular afterload) or similar pulmonary arteryparameters (right ventricular afterload).

The term “preload” refers to the stretching of a single cardiac myocyteimmediately prior to contraction and is therefore related to thesarcomere length.

Since sarcomere length cannot be determined in the intact heart, otherindices of preload such as ventricular end diastolic volume or pressureare used. As an example, preload increases when venous return isincreased.

The term “cardiac myocyte” means a cardiac muscle cell.

The term “stroke volume” (SV) means the volume of blood ejected by theright/left ventricle in a single contraction. It is the differencebetween the end diastolic volume (EDV) and the end systolic volume(ESV). Mathematically, SV=EDV−ESV. The stroke volume is affected bychanges in preload, afterload and inotropy (contractility). In normalhearts, the SV is not strongly influenced by afterload whereas infailing hearts, the SV is highly sensitive to afterload changes.

The term “stroke work” (SW) refers to the work performed by the left orright ventricle to eject the stroke volume into the aorta or pulmonaryartery, respectively. The area enclosed by the pressure/volume loop is ameasure of the ventricular stroke work, which is a product of the strokevolume and the mean aortic or pulmonary artery pressure (afterload),depending on whether one is considering the left or the right ventricle.

The term “ejection fraction” (EF) means the fraction of end diastolicvolume that is ejected out of the ventricle during each contraction.Mathematically, EF=SV/EDV. Healthy ventricles typically have ejectionfractions greater than 0.55. Low EF usually indicates systolicdysfunction and severe heart failure can result in EF lower than 0.2. EFis also used as a clinical indicator of the inotropy (contractility) ofthe heart. Increasing inotropy leads to an increase in EF, whiledecreasing inotropy decreases EF.

The term “end systolic pressure volume relationship” (ESPVR) describesthe maximal pressure that can be developed by the left ventricle at anygiven left ventricular volume, or alternatively, by the right ventricleat any given right ventricular volume. This implies that the PV loopcannot cross over the line defining ESPVR for any given contractilestate. The slope of ESPVR (Ees) represents the end-systolic elastance,which provides an index of myocardial contractility. The ESPVR isrelatively insensitive to changes in preload, afterload and heart rate.This makes it an improved index of systolic function over otherhemodynamic parameters like ejection fraction, cardiac output and strokevolume.

The ESPVR becomes steeper and shifts to the left as inotropy(contractility) increases. The ESPVR becomes flatter and shifts to theright as inotropy decreases.

The term “preload recruitable stroke work relationship” (PRSW) means ameasure of cardiac contractility, and is the linear relationship betweenSW and EDV.

The term “pressure-volume area” (PVA) means the total mechanical energygenerated by ventricular contraction. This is equal to the sum of thestroke work (SW), encompassed within the PV loop, and the elasticpotential energy (PE). Mathematically, PVA=PE+SW.

The term “Langendorff perfusion” refers to a method of perfusing anexcised heart with a nutrient-rich oxygenated solution in a reversefashion via the aorta. The backwards pressure causes the aortic valve toshut thereby forcing the solution into the coronary vessels, whichnormally supply the heart tissue with blood. This feeds nutrients andoxygen to the cardiac muscle, allowing it to continue beating forseveral hours after its removal from the animal.

The term “working heart” as used herein, refers to clinical ex vivocoronary perfusation throughout a excised heart by ventricular fillingvia the left atrium and ejection from the left ventricle via the aortadriven by the heart's contractile function and regular cardiac rhythm.The excised heart is attached by cannulae to a perfusate reservoir andcirculatory pumps in a Langendoff preparation. The flow of perfusatethrough the excised heart in “working heart” mode is in the directionopposite to the flow of perfusate during Langedorff perfusion.

The term “ischemia” means a condition that occurs when blood flow andoxygen are kept from the heart.

The term “conduit” as used herein means cannula.

The present disclosure pertains to apparatus, systems and methods forassessing excised hearts after delivery to transplantation facilities,to determining their suitability for transplantation, and formaintaining selected excised hearts until transplantation.

One exemplary embodiment of the present disclosure relates to anapparatus for receiving, maintaining and assessing an excised donorheart under continuous Langendorff perfusion and periodic “workingheart” perfusion mode. The exemplary apparatus generally comprises ahard-shell container, a removable support for positioning and retainingan excised heart, removable soft-shell reservoir for a perfusate, and alid sealably engagable with the hard shell container. The lid isprovided with a plurality of ports for sealable engaging conduits forinterconnecting: (i) the soft-shell reservoir with a supply of heatedand oxygenated perfusate, (ii) the soft-shell reservoir with an outputpump, (iii) the excised heart with a perfusion cannula inserted into itsaorta, (iv) the excised heart with a perfusion cannula inserted into itspulmonary artery, (v) the excised heart with a perfusion cannulainserted into its pulmonary vein, (vi) the excised heart with aperfusion cannula inserted into its vena cava, and (vii) the excisedheart with a cannula inserted into its vena cava for delivery ofnon-perfusate solutions. The lid may also be provided with ports forprobes and leads for attachment to the cannulae and/or heart.Alternatively, the ports for the probes and leads may be integrallyprovided on a surface of the hard-shell reservoir.

Another exemplary embodiment of the present disclosure relates to asystem for receiving, perfusing and maintaining and assessing an exciseddonor heart. The system generally comprises the above-disclosedapparatus interconnected with: (i) a perfusate-processing system, (ii) abi-directional perfusate pumping system, (iii) flow sensors formonitoring the flow of perfusate to and from an installed heart's aorta,pulmonary artery, pulmonary vein, and vena cava, (iv) an ECG apparatusinterconnectable with the installed heart, and (v) probesinterconnecting the installed heart with instruments for monitoring theheart's physiological functionality using load independent indices andload dependent indices. Suitable perfusate-processing systems areexemplified by heart-lung machines commonly used for coronary bypasssurgeries.

Another exemplary embodiment of the present disclosure relates tomethods for: (i) maintaining an installed excised heart while preservingits physiological functionality by continuous Langendorff perfusion atnormothermic temperatures ranging from about 24° C. to about 35° C., and(ii) assessing the installed excised heart for transplantation underforward perfusion flow using load independent indices exemplified byleft ventricular end systolic pressure volume relationships, rightventricular end systolic pressure volume relationships, left ventricularpreload recruitable stroke work relationships, right ventricular preloadrecruitable stroke work relationships, and isovolumic relaxationconstants measured as Tau. The methods disclosed herein may incorporatemeasurements and assessments of load dependent indices in combinationwith the load independent indices. Suitable load dependent indices areexemplified by diastolic blood pressure, systolic blood pressure, meanblood pressure, among others. The methods disclosed herein for use withthe apparatus and system of the present disclosure can utilize perfusionsolutions known by those skilled in these arts, to be useful forperfusion of excised hearts. Suitable perfusion solutions areexemplified by whole blood, whole blood amended with citrate and/orphosphate and/or dextrose, modified Krebs solutions, Viaspan, modifiedViaspan solutions, and the like.

An exemplary use of the apparatus, system and methods of the presentdisclosure generally comprises the steps of selection, preparation, andbalancing of a perfusate solution, setting up the system byinterconnecting the perfusate-processing system and the bi-directionalperfusate pumping system with cannulae that are subsequentlyinterconnected with the appropriate ports on the lid of the receiving,maintaining, and assessing apparatus, priming the interconnected systemwith the perfusate solution, installing an excised heart onto thesupport provided with the apparatus and then installing the appropriatecannulae into the heart's aorta, pulmonary artery, pulmonary vein, andvena cava, expressing all air from within the heart and the cannulae,and then commencing the Langendorff perfusion at a normothermictemperature from the range of about 25° C. to about 35° C.

It should be noted that the levels of haematocrit, Ca⁺⁺, K⁺, NaHCO₃,Na⁺, pO₂, CO₂, and glucose in the perfusate must be balanced beforeperfusion starts. In the case of using bank CPD donor blood, deranged K⁺and Ca⁺⁺ concentrations may not allow for a homeostatic prime. This canbe adjusted by haemofiltration using Ringers solution as the rinse. Allthese values should ideally start within normal physiological ranges andshould be monitored by inline continuous blood gas analysis. The primarypurpose for the perfusate is to avoid causing tissue edema and tomaintain ion homeostasis to preserve cardiac function.

The bi-directional perfusate pumping system enables the apparatus toprovide Langendorff perfusion flow through isolated aortic root duringthe initial resuscitation, stabilization, and maintenance phases, andthen to switch the direction of perfusate flow to a “working heart” modeduring the pre-transplant assessment phase of the heart's physiologicalfunctionality during which time the preload i.e., atrial pressure can beset at a constant level as also can be the afterload i.e., arterialpressure. The control of these parameters is computer-regulated with avery tight control algorithm to ensure that the heart is notover-distended and therefore avoid potential damage. These pressures canbe precisely controlled to mimic physiological pressure waveforms duringnormal heart cycles.

The present system and methods provide functional assessments of boththe right ventricle and the left ventricle (current systems and methodsonly provide limited functional assessments of the left ventricle). Inaddition to assessing physiological functionality, the present systemand methods provide morphologic assessments through echocardiography andangiography. The combination of morphological assessments withassessments of right ventricle and left ventricle functionality based ona combination of load independent indices and load dependent indicesgreatly improves a clinician's ability to make decisions regarding thesuitability of donor hearts for transplantation. Moreover, this willallow targeted treatment of organ dysfunction such that more hearts canbe recovered and utilized.

The invention includes all embodiments, modifications and variationssubstantially as hereinbefore described and with reference to theexamples and figures. It will be apparent to persons skilled in the artthat a number of variations and modifications can be made withoutdeparting from the scope of the invention as defined in the claimsExamples of such modifications include the substitution of knownequivalents for any aspect of the invention in order to achieve the sameresult in substantially the same way.

Example 1

An exemplary system 10 according to one embodiment of the presentdisclosure is shown in FIGS. 1-3. The system comprises:

-   (1) a perfusion, maintenance, and assessment apparatus comprising a    hard-shell container (not shown) housing: (i) a removal support for    holding an excised heart, (ii) a soft-shell reservoir 20 for    receiving and distributing a perfusate to various entry points in an    excised heart 100, and (iii) a lid 95 that is sealably engagable    with the heard-shell container;-   (2) a perfusate-processing system 30 comprising: (i) a reservoir 31    for receiving egressing perfusate from the perfusion, maintenance,    and assessment apparatus and for receiving fresh perfusate, (ii) a    roller pump 33, and (iii) a heat-exchanger 35 for adjusting    temperature of the perfusate to within the range of about 24° C. to    about 35° C., (iv) an oxygenator (not shown but could be a    stand-alone machine or alternatively, coupled with the heat    exchanger) to condition and balance the perfusate prior to its    conveyance to the soft-shell reservoir 20 in the hard-shell    container;-   (3) a perfusate output pump 40 interconnected with the soft-shell    reservoir 20 and the excised heart 100;-   (4) an ECG machine 90 connectable to the heart 100 with leads 91 and    92 for monitoring the electrical activity of the heart 100;-   (5) monitoring equipment for detecting, recording and/or displaying    and/or transmitting load independent indices data (not shown);-   (6) monitoring equipment for detecting, recording and/or displaying    and/or transmitting load dependent indices data (not shown);-   (7) computer equipment for receiving and processing the load    independent indices data and the load dependent indices data.

The apparatus lid 95 is provided with a plurality of ports for sealableengaging conduits for interconnecting with various entry points into theheart 100 to enable precisely controllable “working heart” perfusion andLangendorff perfusion. The soft-shell reservoir is fitted with threeoutlets to: (i) receive oxygenated and heated perfusate from theperfusate-processing system 30 via conduit 38 (also referred to as acannula); (ii) to preload right atrium via conduits 70 and 76 throughthe vena cava 180 using pressure delivered by the roller pump 33 of theperfusate-processing system 30; and (iii) receive a flow of perfusateejected from the left ventricle through the aorta 150 into conduit 41and then returned to the soft-shell reservoir 20 from the left ventricleunder resistance provided i.e, afterload) by pump 40 into lines 27, 27 aand 21.

Perfusate egressing the soft-shell reservoir 20 through conduit 70 issplit into two flows at juncture 70 a, through conduits 76 and 72, thatare pressurized by roller pump 33. The perfusate flowing through conduit76 and ingressing the right atrium through the vena cava 180 flows intoright ventricle during “working heart mode” perfusion and then throughthe pulmonary artery 170 from where it is conveyed by conduit 36 back tothe perfusate-processing system 30 for conditioning. During Langendorffperfusion, perfusate flow through conduit 72 is directed into conduit 21through juncture 21 a and then pumped into the aorta 150 via conduit 41by centrifugal pump 40. The flow pressure from the soft-shell reservoir20 through conduit 72 is monitored by flow sensor 71, while the flowpressure from the right ventricle into conduit 36 for return to theperfusate-processing system 30 is monitored by flow sensor 37.Integrated pressure ports 28, 42 and 74 monitor and control perfusatepressures at the connection of conduit 21 with the centrifugal pump(i.e., afterload pump) 40, at the connection of conduit 41 with theaorta 150, and at the connection of the line 73 with the pulmonary vein160, respectively. Clamp 26 on conduit 72 is open during Langendorffperfusion while clamp 25 on line 21 is closed thereby causing theperfusate to be supplied only to the aorta 150 and aortic root,providing coronary blood flow, the effluent emerging from the coronarysinus, collected in the right atrium and returned through the rightventricle and pulmonary artery 170 to the hardshell reservoir 31 throughconduit 36. In Langendorff mode, coronary blood flow can be estimatedthrough the flow sensor 37 on conduit 36. During “working heart mode”perfusion, clamp 26 on conduit 72 is closed while clamp 25 on line 21 isopen thereby causing the perfusate to cycle from conduit 27 throughjuncture 72 a into conduit 73 for deliver into the left atrium throughthe pulmonary vein from where in flows into the left ventricle andegresses through the aorta 150 into line 41 and is pumped by the heartto the soft-shell reservoir through conduit 21 with the centrifugal pump40 supplying tightly regulated afterload.

The three pressure sensors 28, 42, 74, the two flow meters 37 and 71,along with the ECG trigger signals augment the computer automationcontrol system for adjusting pump motor speeds, clamp positioning,perfusion system operation, and test modes for the left ventricle. Theautomation system includes motor speed controllers with feedback, sensorreadout hardware and system component synchronization via a personalcomputer running the computation algorithms During perfusion andmaintenance of an excised heart, the heart is secured in the stand inthe hard-shell container which is filled with perfusate held at anormothermic temperature to enable for fluoroscopy across the chamberand echocardiography through the saline. Perfusate flows from thepulmonary artery (right ventricle) into the hard-shell reservoir bygravity. Perfusate is roller pumped from this reservoir through anoxygenator with an integrated heater to the soft-shell reservoir(exemplified by Medtronic's Affinity reservoir bag). During Langendorffperfusion and working heart perfusion, the magnetically-coupledcentrifugal pump (exemplified by Medtronic's BXP-80 pump) will provideflow to the coronary arteries or resistance to the aorta, respectively.Monitoring of atrial pressure, aortic pressure and flow through theheart during working mode enables assessment of cardiac function whilevarying resistance to flow (i.e., afterload), as shown in FIG. 4.

Pressure-volume maps of the ventricles are obtained by deploying apressure wire in a selected ventricle via tubing access ports in thetubing external to the perfusion chamber. Aortic pressure and out-flowprofiles are monitored with an additional flow sensor placed on theaortic output conduit 41 to derive a pseudo pressure-volume curve lessinvasively and on a continuous manner. Similarly, a flow meter placed onthe right atrium inflow conduit 76 and a pressure sensor placed onto theright ventricle output conduit 36 enables recording of continuous pseudopressure-volume graphs.

Echocardiography of the heart is performed through the perfusate-filledperfusion chamber. If required, a block of malleable acoustic-matchinggel is matched to the radius of the perfusion chamber to allow foracoustical coupling. There is a risk that sound wave conduction acrossthe polycarbonate perfusion chamber may be insufficient to allow forimaging. Mitigation of this risk may include thinner chamber wall,different wall material or further conduction enhancement. Angiographyis performed by placing a guide catheter down the aortic cannula andengaging the ostium of the coronary artery under fluoroscopic guidance.During fluoroscopy, contrast is injected down the guide catheter andperfusate is wasted from the system prior to entering the hard-shellreservoir if the volume of contrast would overload the perfusion system.

Example 2

The study disclosed in this example was conducted at the Institute forBiodiagnostics (NRC, Winnipeg, MB, Canada). The protocol was approved bythe local Institutional Animal Care Committee adhering to the guidelinesset by the Canadian Council on Animal Care.

Five domestic swine weighing 40-45 kg were obtained from a commercialsource and acclimatized in the animal housing facility for one weekprior to the study. On the study days, the animals were premedicatedintramuscularly, using atropine 0.05 mg/kg, Midazolam 0.4 mg/kg andKetamine 20 mg/kg Immediately following sedation, general anesthesia wasinduced using 5% Isoflurane in oxygen via a mask. Xylocaine endotrachealspray was applied to the larynx and swine were endotracheally intubatedwith 7.5-mm or 8-mm Portex® tubes (Portex is a registered trademark ofSmiths Medical ASD Inc., Keane, N.H., USA) and mechanically ventilated.An IV catheter was placed in an ear vein and supplemental fluid (0.9%saline) was administered. During surgery, animals were maintained with2.0-2.5% isoflurane. Continuous physiological parameters including, ECG,BP, and end-tidal CO₂ were monitored (PM-9000 Veterinary PortableMonitor, Mindray Corp, Nanshan, China) and kept within acceptablelimits.

Each swine's chest was entered through a midline sternotomy and theheart prepared for retrieval. 300 u/kg heparin was administered at thebeginning of the procedure. Donation after circulatory arrest wassimulated through initiation of hypoxic cardiac arrest via cessation ofmechanical ventilation. Once the animal arrested, a 15-min stand-offperiod was observed. Initial myocardial protection was then achieved byinfusing 250 ml Plegisol® (Plegisol is a registered trademark of HospiraInc., Lake Forest, Ill., USA), 4 ml of KCl (1M) and 250 ml autologousblood into the isolated aortic root at 8° C. 50 mg lignocaine were addedto the reservoir to prevent fibrillation once ex vivo perfusion wasinitiated. Long sections of aorta, pulmonary artery and superior venacava were excised with the heart to ensure adequate space forcannulation with the ex vivo perfusion system. The heart was weighedimmediately after removal from the body. The swine was exsanguinated viathe thoracic aorta, and 1600 ml of blood was returned to the reservoirof the perfusion system by the pump sucker.

After excision, a flexible cone cannula was sewn to the left atrium anda Xvivo® arterial cannula (Xvivo is a registered trademark of XvivoPerfusion AB, Gothenburg, Sweden) was inserted into the ascending aorta.Cannulae with ⅜″ connectors were secured to each of the superior venacava, pulmonary artery, the inferior vena cava, and were oversewn. Theheart was transferred onto the perfusion rig where all cannulae weresecured to ⅜″ PVC tubing. The heart was suspended within a large funnelwith an attached upper splashguard. Securing clamps attached to the mastof the rig stabilized all lines, leaving the heart unrestricted on allsides (FIGS. 2, 3). Biomedicus flow probes (Bio-Probe DP38, MedtronicInc., Minneapolis, Minn., USA) were inserted in the left atrium line andright atrium line.

The saline prime was displaced with swine blood to leave a blood primewith a haematocrit of 20-24%. Sodium bicarbonate and glucose were addedto bring the prime within normal physiological blood ranges. Aftercannulation, the heart was de-aired by atrial filling from the venousbag and an air-free connection was made to the circuit. Perfusion wasstarted after an average 18±6 minutes from the time of cardioplegiadelivery in the donor. Retrograde Langendorff perfusion was commenced at300-500 ml/min with a pressure of 50-60 mmHg from the centrifugal pumpto achieve the coronary blood flow. The venous blood from the coronarysinus was returned to the hard-shell venous reservoir via the pulmonaryartery line. An air inlet line was inserted into the pulmonary arteryvia a ⅜″ Y-connector to prevent a siphon effect. Sweep gas flow wasadjusted to a FiO₂=0.6, 100-200 ml/min O₂ and 10-15 ml/min CO₂ achievedsufficient gas exchange Left atrial, right atrial, aortic and pulmonaryarterial pressures were continuously monitored by recording diastolicblood pressure (dbp), left arterial pressure (lap), flow rate ofperfusate into the left atrium (flow), systolic blood pressure (sbp),and mean blood pressure (mbp). If ventricular fibrillation was required,the heart was defibrillated with 10 joules.

After 15 min of stable Langendorff perfusion, biventricular preload wasinstituted gradually. As the heart started to eject against theretrograde flow the centrifugal pump speed was reduced to 100-200 rpm,which provided afterload to the ejecting left ventricle (FIG. 4). Aspreload was increased by releasing the roller clamps, biventricularoutput increased and afterload was adjusted to provide diastolicpressure (FIG. 4). When measurements with the Biomedicus flow probesindicated that the heart was ejecting, a Ventri-Cath multisegment8-electrode combined pressure/volume catheter (Millar Instruments Inc.,Houston, Tex., USA) was inserted along the longitudinal axis of the leftventricle with the proximal electrode at the level of the aortic valve.In similar fashion, another catheter was inserted into the rightventricle via the pulmonary artery. Data was collected at a samplingrate of 200 Hz with Lab chart 7 (AD Instruments, Bella Vista, NSW,Australia), using a Powerlab AD module. A family of pressure/volumeloops was obtained by manually compressing the left atrial inflow or theright atrial inflow. Subsequently, a dobutamine infusion was started at2.5 ug/Kg/min to 5 ug/kg/min) to allow assessment of contractile reserveand a new set of pressure/volume loops was generated in the same way asbefore (FIG. 7).

During 5 h of perfusion, the physiological variables of the perfusatewere kept within normal limits. During the working mode, the leftventricular end systolic pressure volume relationship (LV ESPVR) was23.1+/−11.1 and the left ventricular preload recruitable stroke workrelationship (LV PRSW) was 67.8+/−7.2 (FIG. 6). Both measures increasedsignificantly after infusion of dobutamine. The LV ESPVR increased to45.1+/−12.2, p=0.01, while the LV PRSW increased to 102.5+/−18.2, p=0.03(FIG. 7). The right ventricular end systolic pressure volumerelationship (RV ESPVR) and the right ventricular preload recruitablestroke work relationship (RV PRSW) were also measured in 2 animals (FIG.8). The RV ESPVR increased from 4.1+/−2.3 to 11.9+/−4.1 after dobutamineinfusion, while the RV PRSW increased from 6.7+/−2.4 to 8.7+/−4.6 afterdobutamine infusion (FIG. 9).

The resuscitated hearts displayed significant upward and leftward shiftsof their LV ESPVR and RV ESPVR, significant increases in their LV PRSWand RV PRSW, and minimal stiffness. Two hearts regained sinus rhythmafter reperfusion, whilst the remaining three required defibrillation.Small boluses of sodium bicarbonate, and glucose were the only drugsadded by the research technician.

The circuit was effective during reperfusion and working modes whilstproving to be successful in maintaining cardiac function in excess offive hours. The use of the Biomedicus pump in delivering Langendorffflow and providing afterload to the working heart was simple andeffective. Diastolic pressures could be effectively manipulated withinstant cardiac response to increased coronary perfusion in the workingphase. The venous bag provided suitable and effective preloads to theleft atrium and the right atrium, and these could be easily adjustedwhen either atria started to stretch.

The conductance catheter was simply inserted via purse strings aroundthe innominate artery and PA, but on several occasions became restrictedby the ventricular trabeculae. Nevertheless, shapes of the captured PVloops were outstanding in this model. There was no evidence of thrombiin the circuits at washout. Blood gases stayed very stable with onlysmall additions of glucose and sodium bicarbonate without any noticeablerise in serum lactate.

After 5 h, the heart was removed from the rig, weighed, dissected andbiopsies were taken. Standard PVAN analysis software (MillarInstruments, Houston, Tex., USA) was used for data analysis (FIGS. 10,11).

Example 3

An exemplary system 200 according to an embodiment of the presentdisclosure for maintenance and monitoring of harvested organs is shownin FIG. 12. The harvested organ maintenance and monitoring system 200comprises:

-   (1) a organ maintenance and perfusion apparatus comprising a    hard-shell container 220 housing a removal support (not shown) for    mounting thereon an excised heart 300, and a lid that is sealably    engagable with the hard-shell container 220. The hard-shell    container 220 also serves as a reservoir for a perfusate solution to    enable total immersion, bathing and perfusion of an excised heart    300 mounted into the removable support;-   (2) a perfusate processing system 230 comprising: (i) a pump 233 for    circulating the perfusate solution from the hard-shell container 220    through (ii) a heat-exchanger 234 for adjusting temperature of the    perfusate to within the range of about 24° C. to about 35° C.,    and (iii) an oxygenator 235 to condition and balance oxygen content    of the perfusate solution prior to its conveyance back into to the    hard-shell container 220;-   (3) a perfusate preloading pump 240 interconnected to the hard-shell    container 220 for providing a flow of the perfusate solution into    the left atrium and the right atrium delivered through an integrated    pressure port 284 and then split into two lines 287, 288 by a “Y”    connecter 286, with line 87 delivering the perfusate solution into    the right atrium and line 288 delivering the perfusate solution into    the left atrium;-   (4) a perfusate afterloading pump 250 interconnected to the    hard-shell container 220 for providing a flow of the perfusate    solution into the aorta 350;-   (5) an ECG machine 290 connectable to the heart 300 with leads 291    and 292 for monitoring the electrical activity of the heart 300;-   (6) monitoring equipment for detecting, recording and/or displaying    and/or transmitting load independent indices data (not shown);-   (7) monitoring equipment for detecting, recording and/or displaying    and/or transmitting load dependent indices data (not shown);-   (8) computer equipment (not shown) for receiving and processing the    load independent indices data and the load dependent indices data.

The apparatus lid is provided with a plurality of ports for sealablyengaging conduits for interconnecting with various entry points into theheart 300 to enable precisely controllable “working heart” perfusion andLangendorff perfusion.

The hard-shell container 220 is provided with at least four ports. Twoof the ports are interconnected to a conduit infrastructure forconveyance of a flow of the perfusate solution from the hard-shellcontainer 220 through the heat-exchanger 234 and oxygenator 235 and thenback into the hard-shell container 220 by pressure provided by pump 233.

Another of the ports interconnects the hard-shell container 220 with theperfusate preloading pump 240. The perfusate preloading pump 240provides a controllable pressurized flow of perfusate solution from thehard-shell container 220 into the right atrium via line 287 left atriumvia line 288. The integrated pressure port 284 interposed the perfusatepreloading pump 240 and the Y connecter 286, communicates with andcooperates with the computer equipment for controllably regulatingpressurized output of perfusate solution from the perfusate preloadingpump 240.

A flow sensor 275 is interposed line 287 to enable electronic monitoringof the flow rate of the perfusate solution delivered from the perfusatepreloading pump 240. A servo-actuated partial occlusion clamp 276 isinterposed in line 287 between the flow sensor 275 and the Y connector286 for regulating perfusate solution flow and pressure delivered intothe right atrium. A pressure port 277 is interposed the flow sensor 275and the right atrium to monitor and communicate the flow rate and/orpressure of perfusate solution delivered into the right atrium by line287. The flow sensor 275, clamp 276, and pressure port 277 communicatewith and cooperate with the computer equipment for adjusting andcontrolling the flow rates of the perfusate solution from the perfusatepreloading pump 240 into the right atrium.

A flow sensor 244 is interposed line 288 to enable electronic monitoringof the flow rate of the perfusate solution delivered from the perfusatepreloading pump 240. A servo-actuated partial occlusion clamp 226 isinterposed in line 288 between the flow sensor 244 and the Y connector286 for regulating perfusate solution flow and pressure delivered intothe left atrium. A pressure port 274 is interposed the flow sensor 244and the left atrium to monitor and communicate the flow rate and/orpressure of perfusate solution delivered into the left atrium by line288. The flow sensor 244, clamp 226, and pressure port 274 communicatewith and cooperate with the computer equipment for adjusting andcontrolling the flow rates of the perfusate solution from the perfusatepreloading pump 250 into the left atrium.

Another of the ports interconnects the hard-shell container 220 with aconduit infrastructure that is connectable to the aorta 350 of the heart300 for maintenance of, monitoring of, and assessing of the functioningof the left ventricle of the heart 300. Pressurized flow of perfusatesolution through this conduit infrastructure is controllably regulatedby an after-loading pump 250, a flow sensor 271, and a pressure port 242interposed the hard-shell container 220 and the aorta 350. Theafter-loading pump 250, the flow sensor 271, and the pressure port 242communicate with, and cooperate with the computer equipment foradjusting and controlling the flow rates of the perfusate solutionflowing through the conduit infrastructure.

During “working heart mode” perfusion, the perfusate solution is pumpedinto the heart 300 by the preloading pump 240 via lines 287, 288 intothe right atrium and the left atrium respectfully. The perfusatesolution is then transferred into the right ventricle 340 and thenegresses through the pulmonary artery 360 through conduit 236 back intothe reservoir of the hard-shell container 220. The flow rate andpressure of the perfusate solution through pressure port 238 intoconduit 236 wherein the flow rate and pressure of the perfusate solutionis monitored by flow meter 237 and regulated by clamp 239. The perfusatesolution ingressing the left atrium via line 288 is transferred into theleft ventricle 320 and then egresses through the aorta 350 into theconduit infrastructure through pressure port 242, flow sensor 271,after-loading pump 250 and then into the hard-shell container 220.

During Langendorff perfusion, the flow of perfusate solution is directedfrom the hard-shell container 220 into the conduit infrastructurethrough after-loading pump 250, flow sensor 271, pressure port 242, andthen into the aorta and the aortic root. The perfusate solution effluentflows through the coronary sinus into the right atrium, then into theright ventricle and eggresses through the pulmonary artery 360 to thehard-shell container 220 through conduit 236. In Langendorff mode,coronary blood flow can be estimated through the flow sensor 237 onconduit 236.

The four pressure sensors 275, 244, 237, and 271, the three clamps 226,276, along with the ECG⁻ trigger signals augment the computer automationcontrol system for adjusting pump motor speeds, clamp positioning,perfusion system operation, and test modes for the left ventricle. Theautomation system includes motor speed controllers with feedback, sensorreadout hardware and system component synchronization via a personalcomputer running the computation algorithms. During perfusion andmaintenance of an excised heart 300, the heart is secured in the stand(not shown) in the hard-shell container 220 which is filled withperfusate solution held at a normothermic temperature to enable forfluoroscopy across the chamber and echocardiography through the saline.Perfusate solution flows from the pulmonary artery (right ventricle)into the hard-shell reservoir under pressure provided by the perfusatepreloading pump 240. The perfusate solution is pumped from thehard-shell container 220 by pump 233 through a heat-exchanger 234 and anoxygenator 235 and then back to the hard-shell container 220.

Pressure-volume maps of the ventricles are obtained by deploying apressure wire in a selected ventricle via tubing access ports in thetubing external to the perfusion chamber. Aortic pressure and out-flowprofiles are monitored with an additional flow sensor placed on theaortic output conduit 41 to derive a pseudo pressure-volume curve lessinvasively and on a continuous manner. Similarly, a flow meter placed onthe right atrium inflow conduit 76 and a pressure sensor placed onto theright ventricle output conduit 36 enables recording of continuous pseudopressure-volume graphs.

Echocardiography of the heart is performed through the perfusate-filledperfusion chamber. If required, a block of malleable acoustic-matchinggel is matched to the radius of the perfusion chamber to allow foracoustical coupling.

There is a risk that sound wave conduction across the polycarbonateperfusion chamber may be insufficient to allow for imaging. Mitigationof this risk may include thinner chamber wall, different wall materialor further conduction enhancement. Angiography is performed by placing aguide catheter down the aortic cannula and engaging the ostium of thecoronary artery under fluoroscopic guidance. During fluoroscopy,contrast is injected down the guide catheter and perfusate is wastedfrom the system prior to entering the hard-shell reservoir if the volumeof contrast would overload the perfusion system.

1-10. (canceled)
 11. An apparatus for perfusing excised donor hearts,comprising: a support for supporting an excised donor heart; a pluralityof conduits for connecting a reservoir of a perfusate to the heartsupported on the support to circulate the perfusate through the heart,said conduits comprising a first cannula for cannulation into an atriumof the heart, and a second cannula for cannulation into an aorta of theheart; a servo-actuated partial occlusion clamp connected to the firstcannula for automated control of a fluid flow through the first cannula;an afterload pump in fluid communication with said second cannula, saidafterload pump configured and operable to selectively apply an afterloadpressure to the aorta to resist circulation of the perfusate through theheart when said clamp is opened to allow the perfusate to enter theatrium through said first cannula; and a controller in electroniccommunication with said clamp and said afterload pump, and configured toautomatically control operation of said clamp and said afterload pumpfor regulating circulation of the perfusate through the heart.
 12. Theapparatus of claim 11, wherein said pump is a centrifugal pump.
 13. Theapparatus of claim 11, wherein the pump is a first pump, and saidapparatus comprises a second pump in fluid communication with the atriumthrough said clamp, for applying a fluid pressure to the atrium.
 14. Theapparatus of claim 13, wherein said second pump is a centrifugal pump.15. The apparatus of claim 11, wherein said atrium is the right atriumof the heart.
 16. The apparatus of claim 11, wherein said controllercomprises a microprocessor.
 17. The apparatus of claim 11, comprising aplurality of flow regulating devices positioned for adjusting perfusateflows in selected ones of said conduits.
 18. The apparatus of claim 11,comprising a temperature sensor positioned for detecting a temperatureof the perfusate, and comprising a heat-exchanger for adjusting thetemperature of the perfusate.
 19. The apparatus of claim 11, comprisingflow sensors positioned for detecting flow rates in selected ones ofsaid conduits.
 20. The apparatus of claim 11, comprising a firstpressure sensor positioned for detecting the afterload pressure, and asecond pressure sensor positioned for detecting the preload pressure.21. The apparatus of claim 11, comprising an oxygenator for controllablyadjusting an oxygen content of the perfusate.
 22. The apparatus of claim11, comprising said reservoir.
 23. The apparatus of claim 11, comprisingan electrocardiographic device.
 24. The apparatus of claim 11,comprising a hard-shell container sealingly engagable with thereservoir, wherein said support is removably mounted in the hard-shellcontainer.
 25. The apparatus of claim 24, wherein the reservoircomprises a soft-shell reservoir housed in said hard-shell container,said soft-shell reservoir comprising ports for sealingly engaging andcommunicating with selected ones of said plurality of conduits.
 26. Theapparatus of claim 24, comprising a lid sealably engagable with thehard-shell container, said lid comprising ports for sealably engagingsaid first cannula and said second cannula.
 27. The apparatus of claim11, wherein said plurality of conduits comprise a resilient conduitconnected to said first cannula, and said clamp clamps around saidresilient conduit for regulating flow of the perfusate through saidresilient conduit.
 28. The apparatus of claim 11, wherein said pluralityof conduits comprises a cannula for cannulation into a pulmonary vein ofthe heart.
 29. A perfusion apparatus for pre-transplant maintenance andassessment of an excised donor heart, comprising: a support forpositioning and mounting thereon the excised heart; a plurality ofconduits for communication with a supply of a perfusate solution, aventricle of the heart, an atrium of the heart, and the aorta of theheart, wherein the plurality conduits comprises a first conduitinfrastructure for conveying a first portion of the perfusate solutionto the atrium; a second conduit infrastructure for conveying a secondportion of the perfusate solution to or from the aorta of the heart; athird conduit infrastructure for conveying a third portion of theperfusate solution from a pulmonary vein of the heart, each one of saidfirst, second, and third conduit infrastructures in communication with apump, a flow sensor, and a controllably adjustable clamp for regulatinga flow of the perfusate solution therethrough.
 30. The perfusionapparatus of claim 29, wherein the controllably adjustable clamp is aservo-actuated partial occlusion clamp.